Selective determination of isoniazid using bentonite clay modified electrodes Uday Pratap Azad, Nandlal Prajapati, Vellaichamy Ganesan PII: DOI: Reference:

S1567-5394(14)00127-3 doi: 10.1016/j.bioelechem.2014.08.011 BIOJEC 6781

To appear in:

Bioelectrochemistry

Received date: Revised date: Accepted date:

21 January 2014 9 August 2014 13 August 2014

Please cite this article as: Uday Pratap Azad, Nandlal Prajapati, Vellaichamy Ganesan, Selective determination of isoniazid using bentonite clay modified electrodes, Bioelectrochemistry (2014), doi: 10.1016/j.bioelechem.2014.08.011

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ACCEPTED MANUSCRIPT Selective determination of isoniazid using bentonite clay modified electrodes Uday Pratap Azad, Nandlal Prajapati and Vellaichamy Ganesan*

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Department of Chemistry, Faculty of Science, Banaras Hindu University, Varanasi-221 005, UP, India Phone: + 91-542-6701609, Fax: + 91-542-2368127 Email: [email protected] and [email protected] * Corresponding author

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Abstract

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Fe(dmbpy)32+ (where dmbpy is 4,4'-dimethyl-2,2'-bipyridine) was immobilized by ion-exchange in a bentonite clay film coating on a glassy carbon electrode. Cyclic voltammetry characteristics of the immobilized Fe(dmbpy)32+ was stable and reproducible corresponding to the Fe(dmbpy)32+/3+ redox process. In the presence of isoniazid (IZ), the electrogenerated in film Fe(dmbpy)33+ oxidized IZ efficiently producing large anodic current. This current was linearly proportional to the IZ concentration in the solution. The process was described by an EC' electrocatalysis mechanism allowing for sensitive determination of IZ with a wide linear dynamic concentration range of 10.0 µM to 10.0 mM. The electrode was tested for its analytical suitability and possible discrimination of interferences by determining IZ in a commercially available pharmaceutical product. The paper reports on a simple, cheap, and easy to fabricate chronoamperometric chemical sensor for determination of IZ. Kinetic parameters, such as the catalytic rate constant (2.3 × 103 M-1 s-1) and diffusion coefficient of IZ (5.42 × 10-5 cm2 s-1), were determined using CV, chronoamperometry, and chronocoulometry. Key-words: Clay modified electrode; Electrocatalysis; Isoniazid oxidation; Bentonite.

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ACCEPTED MANUSCRIPT 1. Introduction

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Regular and/or high intake of isoniazid (IZ), frequently used for treatment of tuberculosis may lead to severe liver problems [1]. As per World Health Organization (WHO) recommendation, a daily dose of IZ should be 4-6 mg/kg body weight [2]. Metabolism of IZ produces hydrazine which induces hyper-toxicity and sometimes may cause death too. Therefore, the patients on this medicine are generally examined for the IZ level at regular intervals. Thus, from the clinical point of view it is necessary to have a sensitive analytical method for real-time quantification of IZ. It is also essential for pharmaceutical industries to determine the IZ level in their products for quality control purposes. Whereas the current methods like high performance liquid chromatography, spectrophotometry, volumetric titrimetry, chemiluminescence, capillary electrophoresis [3-7] require lengthy sample pretreatment procedures for the quantification of IZ, electrochemical methods offer simple and green routes to determine traces of IZ selectively. However, electrochemical oxidation of IZ at a bare glassy carbon (GC) electrode suffers from electrode fouling, sluggish electron transfer kinetics and large oxidation overpotential. Therefore, an electrode with no or negligible electrode fouling property and low oxidation overpotential as well is necessary for the sensitive determination of IZ. Coating the surface of GC with a film of appropriate modifier which can accommodate/incorporate suitable electrocatalyst can achieve this goal. We have reported recently our work on the development of electrochemical sensors for nitrite, IZ and arsenite based on metal complexes of iron immobilized electrodes [8-11]. There is a wide range of modifiers (organic polymers and inorganic matrixes with or without ion exchange groups) which can be used for the purpose [8-25]. However, selection of an electrode modifier depends on the compatibility of the modifier with the electrocatalyst and the analyte (i.e. IZ). Our previous report [9] with Nafion® (Nf) modified electrodes showed slow diffusion of IZ leading to low sensitivity and high detection limit. In this work, a cheap material, clay is used to achieve faster electron transfer with high sensitivity and low detection limit (vide infra). Clays are hydrated aluminum silicate minerals possessing nano-scaled layered structure and large number of ion-exchange sites. The nature and number of ion-exchange sites depends on several factors like interlayer space, the layer charge and location of the layer charge. Cation exchange at these clay minerals depends on the nature and number of ion-exchange sites and chemical composition of solution in contact with the clay mineral. Because of high ion-exchange capacity, strong mechanical stability and large surface area, clays have been widely used as electrode modifiers. The immobilization of transition metal complex cations on clay films as electrocatalysts is a promising approach in the development of electrochemical sensors [11, 22]. In this work we utilized clay as electrode modifier and Fe(dmbpy)32+ (where dmbpy= 4,4’dimethyl-2,2’-bipyridine) as electrocatalyst for selective IZ determination. 2. Experimental 2.1

Reagents and chemicals

Tris(4,4'-dimethyl-2,2'-bipyridine)iron(II) sulphate ([Fe(dmbpy)3]SO4) was synthesized and characterized (λmax = 528 nm, ε = 6411 L mol-1 cm-1) according to the standard literature procedure [26]. Sodium sulphate and poly(vinyl alcohol) (PVA) were purchased from S.D. Fine 2

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chemicals, India. Isoniazid (IZ), bentonite clay (bt) and other chemicals were purchased from Himedia or Qualigens, India. 4,4’-Dimethyl-2,2’-bipyridine was purchased from Sigma Aldrich and used as received. Triply distilled water was used for preparing the aqueous solutions. PVA stock solution (0.1 %) was prepared in hot water with vigorous stirring. Sodium ion-exchanged bt material (Na+-bt) was prepared according to our previous procedure [11]. Electrode fabrication

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Aqueous colloid of Na+-bt (0.1%) was prepared together with 0.01% of PVA in water. A 20 µL sample of this colloid was dropped onto the surface of GC and dried at room temperature at least for 6 h. The dried electrode was dipped in triply distilled water for 10 min (represented as GC/Na+-bt) before the immobilization of Fe(dmbpy)32+ by ion-exchange process. In the next step, the GC/Na+-bt was dipped into aqueous 0.5 mM Fe(dmbpy)32+ solution for 2 h. Long dipping time (2 h) was used in order to ion-exchange/adsorb maximum amount of Fe(dmbpy)32+ into/onto the Na+-bt film. After 2 h, the electrode was removed from Fe(dmbpy)32+ solution and rinsed with triply distilled water in order to remove any physically adsorbed excess Fe(dmbpy)32+. In the last step, the electrode was placed in 0.1 M Na2SO4 solution for a few min. Then, the electrode was transferred to the electrochemical cell containing the supporting electrolyte (0.1 M Na2SO4) and continuously scanned from 0.6 to 1.3 V for 20 times to achieve stable redox peak currents. The electrodes prepared in this fashion are denoted hereafter as GC/Na+-bt/Fe(dmbpy)32+. After obtaining stable redox response, the electrodes were used for electroanalytical applications or kinetic measurements. Commercial fabrication of GC/Na+-bt electrodes would be less expensive than the fabrication of GC/Nf electrodes [9] since clay is quite cheap and commercially available easily as compared to Nf. For absorption spectral studies, Na+-bt and Fe(dmbpy)32+ exchanged Na+-bt films were prepared on clean glass plates similar to the films on the GC electrodes. However, dipping in 0.1 M Na2SO4 solution and potentiodynamic scanning was not carried out. The thickness of the Na+-bt film on the GC electrodes and on the glass plates was kept same by adjusting the amount of colloid used to prepare the films. 2.3 Apparatus

FT-IR spectra of Na+-bt and Fe(dmbpy)32+ exchanged Na+-bt clay material was recorded on PerkinElmer spectrometer (spectrum two, UK) using KBr pellets. Cyclic voltammetry (CV) and other electrochemical measurements were carried out in a one-compartment three-electrode cell. GC/Na+-bt or GC/Na+-bt/Fe(dmbpy)32+ was used as working electrode with an auxiliary electrode of platinum wire and a reference, saturated calomel electrode (SCE). All the experiments were carried out at room temperature (25 ºC) and triply distilled water was utilized throughout the experiment. 3. Result and discussion 3.1

Materials characterization

To understand the ion-exchange/incorporation and possible interaction of Fe(dmbpy)32+ with Na+-bt material, FT-IR studies were performed. Fig. 1 depicts the FTIR 3

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spectra of Na+-bt and Fe(dmbpy)32+ immobilized Na+-bt materials. IR vibrations characteristic of clay minerals are distinctly exhibited by Na+-bt (Fig. 1a) [11]. For example, Al-OH-Al and SiOH stretching vibrations in Na+-bt are observed by the presence of bands at 3621 and 3696 cm-1. The important characteristic vibrations and their corresponding bands are given in supporting information Table S1 [11,27-30]. In the case of Fe(dmbpy)32+ immobilized Na+-bt material (Fig. 1b) additional bands observed at 1483 and 1428 cm-1 are attributed for C=C stretching vibration whereas the band observed at 834 cm-1 may be assigned to heterocyclic ring deformation (Table S1) [28,29]. The appearance of these bands suggests the successful incorporation of Fe(dmbpy)32+ in Na+-bt material. The less intense bands are attributed to the small amount of Fe(dmbpy)32+ ion-exchanged onto the Na+-bt [31]. Fig. 2 shows the UV-Vis absorption spectra of Fe(dmbpy)32+ exchanged Na+-bt film and

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the aqueous Fe(dmbly)32+ solution. Aqueous Fe(dmbpy)32+ shows absorption peak at 528 nm whereas Fe(dmbpy)32+ immobilized on the Na+-bt film shows the peak at 532 nm. Thus on immobilization, Fe(dmbpy)32+ shows a small (4 nm) red shift. However, the shape of the spectrum is changed considerably. The observed red shift and the change in the shape of the spectrum could be due to strong interaction of Fe(dmbpy)32+ with Na+-bt film. In addition to the possible stacking of Fe(dmbpy)32+ within the interlayer space of clay plates, Fe(dmbpy)32+ is possessed by two ion-exchange sites of clay [32,33] which makes them to bind strongly with clay. 3.2 Electrochemical characterization of Fe(dmbpy)32+ in clay film After preliminary scans (refer section 2.2), the stable cyclic voltammograms of GC/Na+bt/Fe(dmbpy)32+ electrode in 0.1 M Na2SO4 recorded at different scan rates are shown in Fig. 3. Under similar conditions, no voltammetric response is observed for Here Fig. 3

GC/Na+-bt (Fig. not shown). However, well defined anodic and cathodic peaks are observed for GC/Na+-bt/Fe(dmbpy)32+ with an E1/2 (E1/2 = (Epa+Epc)/2) value of 0.69 V (scan rate = 20 mVs1 ). The anodic and cathodic peak current increases with successive increase in the scan rate. Anodic to cathodic peak current ratio (Ipa/Ipc) is found to be 0.67 (scan rate = 20 mVs-1) which is lower than the expected value for a perfect reversible system. Anodic to cathodic peak separation, ΔEp (ΔEp = Epa-Epc) is calculated as 86 mV (scan rate = 20 mVs-1) which is higher than the expected value (59 mV) for the one-electron transfer reversible redox reaction. It should be noted that aqueous Fe(dmbpy)32+ exhibits characteristics of a reversible system [34] when unmodified GC is used (supporting information Fig. S1). Increased peak separation and decreased Ipa/Ipc ratio observed in the present system, indicate the poor reversibility of Fe(dmbpy)32+ in clay film [35]. It seems that the oxidation process is slow and some of the Fe(dmbpy)32+ species oxidizes at higher potential than the peak potential without contributing to 4

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the peak current. This kind of strange behavior is not reported at the clay modified electrode and it is probably due to the strong interaction of Fe(dmbpy)32+ species with clay layers and unfavorable stacking/interaction of the oxidized species Fe(dmbpy)33+. On further analysis, the linear variation of Ipa and Ipc with the square root of scan rates (ν1/2), signifies that the redox processes (Fe(dmbpy)32+/3+) in the Na+-bt film is diffusion controlled (inset of Fig. 3). 3.3 Electrocatalytic oxidation and determination of IZ

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The electrocatalytic activity of immobilized Fe(dmbpy)32+ is tested for the oxidation of IZ. It is studied by recording cyclic voltammograms of GC/Na+-bt and GC/Na+-bt/Fe(dmbpy)32+ in absence and presence of 1.0 mM IZ (Fig. 4). In the absence of IZ, GC/Na+-bt shows no characteristic peaks

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(Fig. 4a) whereas GC/Na+-bt/Fe(dmbpy)32+ shows well defined redox peaks due to Fe(dmbpy)32+/3+ redox processes (Fig. 4b) as discussed previously (section 3.2). In the presence of 1.0 mM IZ, the oxidation process of IZ at GC/Na+-bt (Fig. 4a’) is sluggish as evidenced by the broad peak at high overpotential (above 1000 mV). When GC/Na+-bt/Fe(dmbpy)32+ is used for the oxidation of IZ under the similar conditions (Fig. 4b’) a relatively sharp peak al low overpotential (850 mV) is observed with high oxidation current. The corresponding cathodic peak for the reduction of Fe(dmbpy)33+ to Fe(dmbpy)32+ is completely absent demonstrating the efficient reaction between the electrogenerated Fe(dmbpy)33+ with IZ. The remarkable enhancement in the oxidation peak current (about 3 times higher than at GC/Na+-bt at 850 mV), the complete absence of reduction current and significant decrease in the IZ oxidation overpotential (more than 150 mV) are the strong evidences of electrocatalytic effect of the immobilized Fe(dmbpy)32+ species towards the oxidation of IZ. These results clearly indicate the improvement in the electron transfer kinetics of IZ oxidation in presence of Fe(dmbpy)32+ species. This strong electrocatalysis phenomenon [8-11] can be applied for the quantitative determination of IZ. For that, cyclic voltammograms in the presence of increasing concentrations of IZ (from 10.0 µM to 100.0 mM) is analyzed using GC/Na+-bt/Fe(dmbpy)32+ electrode which are shown in Fig. 5. On increasing the concentration of IZ, the resulting anodic current is also increases up to 10.0 mM (inset of Fig. 5). Here Fig. 5 Above 10.0 mM, the peak current deviates from linear behavior with large fluctuation (Fig. S2), which could be due to the oxidation products of IZ. The oxidation products formed on the surface of the GC/Na+-bt/Fe(dmbpy)32+ electrode may affect the regeneration of the catalyst in the Na+-bt/Fe(dmbpy)32+ film. The electrode exhibits a sensitivity of 0.009 (± 0.002) µA/µM and a detection limit of 0.8 (± 0.1) μM (based on 3 times standard deviation of the blank). Thus the present method substantially lowers the oxidation potential, decreases the detection limit and increases the sensitivity than the recent method reported for IZ determination [9]. 3.4 Kinetics and mechanism of IZ oxidation

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ACCEPTED MANUSCRIPT Since the GC/Na+-bt/Fe(dmbpy)32+ exhibits consistent and reversible electrochemical properties towards the electrocatalytic oxidation of IZ, the evaluation of catalytic rate constant (kC) can offer useful insights to the electron transfer kinetics. Chronoamperometry can be used for the estimation of kC as per the equation 1 [10,36,37].

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IC/I = (πC0tkC)1/2

(1)

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where Ic and I are the currents observed at the GC/Na+-bt/Fe(dmbpy)32+, in the presence and absence of 1.0 mM IZ, respectively. C0 is the concentration of IZ in the electrochemical cell and t is the time. From the slope of IC/I vs. t1/2 plot (supporting information Fig. S3), kC was estimated to be 2.3 × 103 M-1s-1 which is comparable to reported value [36] for (FcM)TMA catalyst at platinum electrode and indicate the efficient electrocatalytic oxidation. Chronoamperometry experiments are performed at different concentrations of IZ at GC/Na+bt/Fe(dmbpy)32+ electrode to determine the diffusion coefficient (D) of IZ and Cottrell equation is employed for the determination [34] (equation 2).

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I = π−1/2t-1/2nFACD1/2

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Where C is the bulk concentration of IZ, I is the current due to IZ, and other symbols signify their usual meaning. When the scan rate is increased (at a fixed concentration of IZ as 1.0 mM), the IZ oxidation current also increased with a slight shift in the oxidation peak potential to higher values (supporting information Fig. S4). The variation of oxidation current (Ipa) with the square root of scan rate (ν1/2) is linear (insert of supporting information Fig. S4) indicating the oxidation of IZ at GC/Na+-bt/Fe(dmbpy)32+ electrode is a diffusion controlled process. The slight shift in oxidation peak potential towards positive direction with increase in scan rate indicates kinetic limitation between Fe(dmbpy)32+ immobilized in Na+-bt film and IZ [38]. The plot of ν1/2 vs Ipa/ν1/2 depicts exponential decrease like cure for IZ oxidation (supporting information Fig. S5). The shape of the curve indicates the electrocatalytic EC’ mechanism at the GC/Na+bt/Fe(dmbpy)32+ for IZ oxidation [39]. Further to get information about the number of electrons involved in the rate determining step, Tafel plot is drawn. From the slope of the log(Epa) vs log(ν) plot (equation 3) (supporting information Fig. S6) Epa

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value of Tafel slope (b) is calculated as 101 mV suggesting that only one electron is involved in the rate determining step [37,39]. Plot of I vs t-1/2 is drawn for various concentrations of IZ which (Fig. 6) were found to be linear. From the slopes of the resulting linear plots and Here Fig. 6 concentration of IZ, a plot shown in the insert of Fig. 6 is prepared (Slope vs [IZ]). The average value of D is calculated as 5.42 × 10-5 cm2s-1 which is ten times faster than our previous report [9] with Nf and comparable to other reported literature value [40]. The above results together with comparable kC and D values indicate that the efficiency and mechanism of IZ oxidation is 6

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In the last step, chemical decomposition of

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probably same as that of other types of modified electrodes (like at overoxidized polypyrrole modified GC and 4-pyridyl hydroquinone self-assembled modified microdisk platinum electrodes [7,40]). Thus the present study suggests a similar mechanism [9,41,42] for the electrocatalytic oxidation of IZ at GC/Na+-bt/Fe(dmbpy)32+ as shown in equations 4 and 5.

3.5 Determination of IZ in a pharmaceutical tablet

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The GC/Na+-bt/Fe(dmbpy)32+ electrodes are used to determine the amount of IZ present in real samples. Determination of IZ selectively in the presence of another bacteriostatic drug, rifampin is a difficult task yet essentially required in the quality control units of pharmaceutical industries. Therefore, a pharmaceutical formulation (tablet) containing rifampin (450 mg) and IZ (300 mg) is chosen for demonstration and pertinence of this electrode for the determination of IZ. The calculated amount of powdered tablet is fully dissolved in water and analyzed for its IZ content at GC/Na+-bt/Fe(dmbpy)32+. Using the previously constructed calibration plot (inset of Fig. 5) and the measured oxidation current for the tablet solution, the concentration of IZ is evaluated. Thus estimated IZ concentration agrees well with the IZ concentration actually present in the tablet (average of three measurements), indicating no interference from rifampin for the determination IZ. Cyclic voltammograms of IZ tablet solution containing rifampin is shown in supporting information Fig. S7. Though this method is selective for the determination of IZ in presence of rifampin, this present method may not be suitable for the determination of IZ in the presence of nitrite or arsenite [8,10,11,18]. Further to validate this method, recovery analysis was done by the addition of varying amounts of standard IZ. The resulting percentage of recovery is acceptable and found to be in the range 99.7-102.5% (Table 1). Relative standard deviation (RSD) for the three repeated analysis is also shown for Here Table 1 each determination (Table 1) which are in the range 2.2-3.3 %. These results demonstrate the successful determination of IZ present in the pharmaceutical formulation. 3.6 Stability and reproducibility Reproducibility and stability are the most important parameters which must be taken always in consideration for the development of a significant electrochemical sensor. To clarify 7

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precision/reproducibility of the method/electrode for the determination of IZ, electrochemical experiments are repeatedly performed 10 times with a single GC/Na+-bt/Fe(dmbpy)32+ electrode in the solution containing 1.0 mM of IZ. The relative standard deviation (RSD) of the oxidation current is found to be 2.4%, which reveals the good reproducibility of the present electrode for the determination of IZ. Operational stability of the suggested electrode is checked by measuring the oxidation current before and after completion of the experiments. It shows about 1 to 2% decrease in the initial oxidation current of the catalyst with an RSD of 3.4% indicating the acceptable operational stability. The prepared electrodes are stable for long period of time (stored in water and tested for a month based on the oxidation current of the catalyst).

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4. Conclusions

Acknowledgements

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Electrochemical properties of Fe(dmbpy)32+ immobilized on bentonite films are evaluated by cyclic voltammetry, chronoamperometry, and chronocoulometry techniques. These films exhibit stable and reproducible redox current after few initial scans. Further, they show proficient catalytic activity towards the electro oxidation of IZ. This property is consequently utilized to construct an electrochemical sensor for IZ determination. Presence of rifampin does not alter the oxidation current in the determination of IZ. This inexpensive metal complex based sensor shows linear response in a wide calibration range from 10 µM to 10.0 mM. The applicability of the proposed sensor in the determination IZ present in real sample i.e. in a commercially available tablet is demonstrated.

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Generous funds from CSIR (01(2708)/13/EMR-II) and UGC (42-271/2013 (SR)), New Delhi are gratefully acknowledged. We thank Prof. S. K. Sengupta and Dr. P. Adhikary for useful suggestions.

Appendix A. Supplementary data Supporting information as mentioned in the text can be found online at http://dx.doi.org/….

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References S. Shahrokhian, E. Asadiana, Simultaneous voltammetric determination of ascorbic acid, acetaminophen and isoniazid using thionine immobilized multi-walled carbon nanotube modified carbon paste electrode, Electrochim. Acta 55 (2010) 666-672.

[2]

C. Becker, J.B. Dressman, G.L. Amidon, H.E. Junginger, S. Kopp, K.K. Midha, V.P. Shah, S. Stavchansky, D.M. Barends, Biowaiver monographs for immediate release solid oral dosage forms: Isoniazid, J. Pharm. Sci. 96 (2007) 522-531. P.F. Fang, H.L. Caia, H.D. Li, R. H. Zhu, Q.Y. Tana, W. Gao, P. Xua, Y.P. Liu, W.Y. Zhang, Y.C. Chen, F. Zhang, Simultaneous determination of isoniazid, rifampicin, levofloxacin in mouse tissues and plasma by high performance liquid chromatographytandem mass spectrometry, J. Chromatogr. B 878 (2010) 2286-2291. P. Nagaraja, K.C. Srinivasa Murthy, H.S. Yathirajan, Spectrophotometric determination of isoniazid with sodium 1,2-naphthoquinone-4-sulphonate and cetyltrimethyl ammonium bromide, Talanta 43 (1996) 1075-1080. M. Sarwar, A. Malick, U.A. Khan, Determination of isonicotinic acid hydrazide using Nbromosuccinimide, Anal. Lett. 22 (1989) 853-860. S.C. Zhang, H. Li, Flow-injection chemiluminescence sensor for the determination of isoniazid, Anal. Chim. Acta 444 (2001) 287-294. T.Y. You, L. Niu, J.Y. Gui, Detection of hydrazine, methylhydrazine and isoniazid by capillary electrophoresis with a 4-pyridyl hydroquinone self-assembled microdisk platinum electrode, J. Pharma. Biomed. Anal. 19 (1999) 231-237. U.P. Azad, V. Ganesan, Efficient sensing of nitrite by Fe(bpy)32+ immobilized Nafion modified electrodes, Chem. Commun. 46 (2010) 6156-6158. U.P. Azad, V. Ganesan, Efficient electrocatalytic oxidation and selective determination of isoniazid by Fe(tmphen)32+-exchanged Nafion®-modified electrode, J. Solid State Electrochem. 16 (2012) 2907-2911. U.P. Azad, V. Ganesan, Tris(4,4'-dimethyl-2,2'-bipyridine)iron(II) immobilized Nafion® for arsenite determination in water samples, ChemElectroChem 2 (2014) 379-383. U.P. Azad, S. Turllapati, P.K. Rastogi, V. Ganesan, Tris(1,10-phenanthroline)iron(II)bentonite film as efficient electrochemical sensing platform for nitrite determination, Electrochim. Acta 127 (2014) 193-199. V. Ganesan, P.K. Rastogi, R. Gupta, M.T. Meredith, S.D. Minteer, Ion exchange voltammetry at branched polyethylenimine cross-linked with ethylene glycol diglycidyl ether and sensitive determination of ascorbic acid, Electrochim. Acta 105 (2013) 31-39. P.K. Rastogi, V. Ganesan, S. Krishnamoorthi, Ion exchange voltammetry at permselective copolymer modified electrode and its application for the determination of catecholamines, J. Electroanal. Chem. 676 (2012) 13-19. P.K. Rastogi, V. Ganesan, S. Krishnamoorthi, Microwave assisted polymer stabilized synthesis of silver nanoparticles and its application in the degradation of environmental pollutants, Mater. Sci. Eng. B 177 (2012) 456-461. P.K. Rastogi, S. Krishnamoorthi, V. Ganesan, Synthesis, characterization and ion exchange voltammetry study on 2-acrylamido-2-methylpropane sulphonic acid and N(hydroxymethyl) acrylamide-based copolymer, J. Appl. Polym. Sci. 123 (2012) 929-935.

[7]

[8] [9]

[10] [11]

[12]

[13]

[14]

[15]

RI

SC

NU

MA

[6]

D

[5]

TE

[4]

AC CE P

[3]

PT

[1]

9

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[22] [23]

[24]

[25] [26]

[27]

[28] [29] [30]

[31] [32]

[33]

PT

RI

SC

[21]

NU

[20]

MA

[19]

D

[18]

TE

[17]

M. Pal, V. Ganesan, Zinc phthalocyanine and silver/gold nanoparticles immobilized MCM-41 type materials as electrode modifiers, Langmuir 25 (2009) 13264-13272. M. Pal, V. Ganesan, Effect of silver nanoelectrode ensembles on the electrocatalytic reduction of NO2- by zinc phthalocyanine, Electrochim. Acta 55 (2010) 4071-4077. M. Pal, V. Ganesan, Electrochemical determination of nitrite using silver nanoparticles modified electrode, Analyst 135 (2010) 2711-2716. M. Pal, V. Ganesan, Effect of gold nanoparticles on the electrocatalytic reduction of oxygen by silica encapsulated cobalt phthalocyanine, J. Electroanal. Chem. 672 (2012) 711. M. Pal, V. Ganesan, Electrocatalytic activity of cobalt Schiff base complex immobilized silica materials towards oxygen reduction and hydrazine oxidation, Catal. Sci. Technol. 2 (2012) 2383-2388. M. Pal, V. Ganesan, U.P. Azad, Photochemical oxygen reduction by zinc phthalocyanine and silver/gold nanoparticle immobilized silica thin films, Thin Solid Films 525 (2012) 172-176. C. Mousty, Sensors and biosensors based on clay-modified electrodes-new trends, Appl. Clay Sci. 27 (2004) 159-177. M. Satyanarayana, K.K Reddy, K.V. Gobi, Multiwall carbon nanotube ensembled biopolymer electrode for selective determination of isoniazid in vitro, Anal. Methods, 6 (2014) 3772-3778. P.R. Martins, L.M.C. Ferreira, K. Araki, L. Angnes, Influence of cobalt content on nanostructured alpha-phase-nickel hydroxide modified electrodes for electrocatalytic oxidation of isoniazid, Sens. Actuators B 192 (2014) 601-606. B.K. Jena, C.R. Raj, Au nanoparticle decorated silicate network for the amperometric sensing of isoniazid, Talanta 80 (2010) 1653-1656. M.H. Ford-Smit N. Sutin, The kinetics of the reactions of substituted 1,10phenanthroline, 2,2’-dipyridine and 2,2’,2”-tripyridine complexes of iron(III) with iron(II) ions, J. Am. Chem. Soc. 83 (1961) 1830-1834. S. Tunc, O. Duman, The effect of different molecular weight of poly(ethylene glycol) on the electrokinetic and rheological properties of Na-bentonite suspensions, Colloids Surf. A 317 (2008) 93-99. P. Jing, M. Hou, P. Zhao, X. Tang, H. Wan, Adsorption of 2-mercaptobenzothiazole from aqueous solution by organo-bentonite, J. Environ. Sci. 25 (2013) 1139-1144. H. Jiang, W. Sun, R. Zheng, Synthesis and magnetic properties of novel containing 1,10phenanthroline polymeric complexes, Eur. Polym. J. 42 (2006) 425-433. V. Amani, N. Safari, B. Notash, H.R. Khavasi, Synthesis, characterization and crystal structure determination of iron(III) complexes containing 4,4′- dimethyl-2,2′-bipyridine, dimethyl sulfoxide and chloride, [Fe(dmbipy)Cl4] [dmbipyH] and [Fe(dmbipy)Cl3(DMSO)], J. Coord. Chem. 62 (2009) 1939-1950. K. Jindal, M. Tomar, V. Gupta, Nitrogen-doped zinc oxide thin films biosensor for determination of uric acid, Analyst 138 (2013) 4353-4362. T. Eyama, Y. Yogo, T. Fujimura, T. Tsukamoto, D. Masui, T. Shimada, H. Tachibana, H. Inoue, S. Takagi, Adsorption and stacking behaviour of zwitterionic porphyrin on the clay surface, Clay Minerals 47 (2012) 243-250. H. Sato, A. Yamagishi, S. Katos, Monte Carlo Simulations for the Interactions of Metal Complexes with the Silicate Sheets of a Clay: Comparison of Binding States between

AC CE P

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[38]

[39] [40]

PT

AC CE P

[42]

TE

D

[41]

RI

[37]

SC

[36]

NU

[35]

MA

[34]

Tris(1,10 phenanthroline)metal(II) and Tris( 2,2'-bipyridyl)metal(II) Chelates, J. Am. Chem. Soc. 114 (1992) 10933-10940. A.J. Bard, L.R. Faulkner, Electrochemical methods; Fundamentals, Applications, Wiley, New York, 1980. U.P. Azad, V. Ganesan, Influence of metal nanoparticles on the electrocatalytic oxidation of glucose by poly(NiIIteta) modified electrodes, Electroanalysis 22 (2010) 575-583. Z.N. Gao, X.X. Han, H.Q. Yao, B. Liang, W.Y.Liu, Electrochemical oxidation of isoniazid catalyzed by (FcM)TMA at the platinum electrode and its practical analytical application, Anal. Bioanal. Chem. 385 (2006) 1324-1329. U.P. Azad, V. Ganesan, Determination of hydrazine by polyNi(II) complex modified electrodes with a wide linear calibration range, Electrochim. Acta 56 (2011) 5766-5770. S.J.R. Prabakar, S.S. Narayanan, Amperometric determination of hydrazine using a surface modified nickel hexacyanoferrate graphite electrode fabricated following a new approach, J. Electroanal. Chem. 617 (2008) 111-120. S.M. Golabi, H.R. Zare, Electrocatalytic oxidation of hydrazine at a chlorogenic acid (CGA) modified glassy carbon electrode, J. Electroanal. Chem. 465 (1999) 168-176. M.R. Majidi, A. Jouyban, K.A. Zeynali, Voltammetric behavior and determination of isoniazid in pharmaceuticals by using overoxidized polypyrrole glassy carbon modified electrode, J. Electroanal. Chem. 589 (2006) 32-37. K. Johnsson, P.G. Schultz, Mechanistic studies of the oxidation of isoniazid by the catalase peroxidase from mycobacterium tuberculosis, J. Am. Chem. Soc. 116 (1994) 7425-7426. M.S. Frank, P.V.K. Rao, Kinetics of oxidation of nicotinoyl and isonicotinoyl hydrazines by iron(III) in presence of 1,10-phenanthroline, Indian. J. Chem. 17A (1979) 632-634.

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Recovery (%)

RSD (%) (n = 3)

0

92

94.3

PT

102.5

2.2

5

97

100.3

101.0

3.3

10

102

101.7

99.7

2.4

20

112

114.4

102.1

2.9

RI

Amount of IZ found (mg) (average of three measurements)

MA D

92

Total amount present in the sample (mg) (average of three measurements)

TE

IZ tablet

Amount of standard IZ spiked (mg)

NU

Amount of IZ present in tablet (mg)

AC CE P

Sample

Determination of IZ in a pharmaceutical tablet.

SC

Table 1.

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Figure captions

PT

Fig. 1. The FTIR transmission spectra of the (a) Na+-bt and (b) Fe(dmbpy)32+ exchanged Na+-bt materials.

SC

RI

Fig. 2. The UV-vis spectra of aqueous solution of Fe(dmbpy)32+ (a) and Fe(dmbpy)32+ immobilized Na+-bt film (b).

NU

Fig. 3. The CV curves for GC/Na+-bt/Fe(dmbpy)32+ electrode at potential scan rate of (1) 10, (2) 20, (3) 50, (4) 100, (5) 150, (6) 200, (7) 250, (8) 300, (9) 350, (10) 400 and (11) 500 mVs-1 in 0.1 M Na2SO4. Inset shows the corresponding Ipa and Ipc vs. square root of scan rate (ν1/2) plot.

MA

Fig. 4. The CV curves for GC/Na+-bt (a, a’) and GC/Na+-bt/Fe(dmbpy)32+ (b, b’) electrodes in 0.1 M Na2SO4 in absence (a, b) and presence (a’, b’) of 1.0 mM of IZ at a scan rate of 20 mVs-1.

TE

D

Fig. 5. The CV curves for GC/Na+-bt/Fe(dmbpy)32+ electrode in 0.1 M Na2SO4 with (1) 0.01, (2) 0.03, (3) 0.05, (4) 0.1, (5) 0.5, (6) 1.0, (7) 2.0, (8) 2.5, (9) 5.0, (10) 8.0 and (11) 10.0 mM additions of IZ at potential scan rate of 20 mV s-1. Inset shows the linear dependence of the anodic peak current, Ipa/µA, on the IZ/mM concentration in the range of 0.01 to 10.0 mM.

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Fig. 6. The anodic peak current, Ipa/µA, vs. reciprocal of square root of time (t-1/2/s-1/2) plots for the IZ concentration of (a) 0.2, (b) 0.4, (c) 0.6, (d) 0.8 and (e) 1.0 mM. Inset shows the slope/µA s1/2 vs. IZ concentration, [IZ]/mM plot.

13

3000 1428

2000

14

/ cm -1 1000

532

914 790

1638

PT

RI

SC

a

832

1124 1032

MA

NU

3696

3440

b

1483

D

TE

AC CE P

3621

T/%

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Fig. 1

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PT

Fig. 2

AC CE P

TE

D

MA

A

b

NU

SC

RI

a

500

600

 / nm

15

700

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11

RI

PT

2

SC NU

1

MA

0

0.6

1.0

I / 

D 0.4

1.5

Ipa

0.5 0.0

-0.5

TE

-1

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I / 

1

Ipc

-1.0 -1.5 5

10

15

20

1/2 / mV s-1/2

0.8

1.0

1.2

E / V (vs. SCE)

16

1.4

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RI

PT

Fig. 4

SC NU

b

TE

D

4

0

a'

MA

8

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I / 

12

0.4

0.6

b'

a

0.8

1.0

1.2

E / V (vs. SCE)

17

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80

11

SC

60 40

NU

20 0

2

4

MA

0 6

8

CIZ / mM

10

TE

D

50

0

AC CE P

I / A

100

RI

100

I / A

150

PT

Fig. 5

0.4

0.6

1 0.8

1.0

E / V (vs. SCE)

18

1.2

1.4

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Fig. 6

SC

35 30

0.2

0.4

0.6

0.8

1.0

CIZ / mM

d

NU

25 20

e

RI

40

c

MA

40

45

b

D

30

10

a

TE

20

AC CE P

I / 

50

Slope / As

1/2

60

0.6

0.7

t

0.8 -1/2

0.9 -1/2

/s

19

1.0

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PT

Highlights

 Fe(dmbpy)32+ -bentonite film is used for isoniazid (IZ) determination.

RI

 It demonstrates efficient catalytic activity towards the electro oxidation of IZ

SC

 IZ present in a tablet is determined in the presence of rifampin.

AC CE P

TE

D

MA

NU

 The sensor exhibits high selectivity and sensitivity for IZ detection.

20